Membrane proteins constitute about 30 % of all prokaryotic and eukaryotic proteomes. They are responsible for ion conduction, chemical transport, energy conversion, signal transduction, hormone- and photo-reception, cell adhesion, and many other functions. The long-held tenet of structural biology is that to fully understand the function of biomolecules their structure must be known. Membrane proteins are no exception in this regard. However, the science aimed at elucidating their structures has lagged far behind similar science on soluble proteins, mostly for technical reasons. The goal of the research proposed in this application is two-fold: (1) to advance solution NMR spectroscopy as a method for structure determination of membrane proteins and (2) to further the understanding of the functions of four prokaryotic outer membrane proteins by studying their structures by solution NMR. Specifically: OmpA from E. coli primarily will be used as a model membrane protein for NMR methods development, for studying lipid-protein interactions, and for examining the stability of membrane proteins in lipid bilayers;OmpG from E. coli is becoming the preferred biological nanopore for biosensor development and NMR will be used to understand the mechanism, by which it gates from the open to the closed state;and OprG and OprH are two outer membrane proteins from Pseudomonas aeruginosa, whose contributions to the severe antibiotic resistance of this microorganism will be investigated by determining their structures and studying their functions. PUBLIC HEALTH RELEVANCE: This project will push the frontier of determining the structures and understanding the functions of membrane-bound proteins by nuclear magnetic resonance spectroscopy. The specific membrane proteins that will be examined have biotechnological applications in biosensor development or are components of the cell envelope of Pseudomonas aeruginosa, i.e. a clinically important pathogen in hospital-induced infections and complications of cystic fibrosis. The research may ultimately contribute to better treatments of these infections.